Occurrence and partial genetic characterisation of Lettuce big‐vein associated virus and Mirafiori lettuce big‐vein virus infecting lettuce in Jordan

Lettuce big‐vein disease (LBVD) is an economically damaging disease that occurs in most lettuce production areas worldwide. Several studies have associated LBVD with a complex of two viruses, Lettuce big‐vein associated virus (LBVaV) and Mirafiori lettuce big‐vein virus (MiLBVV). Both viruses are transmitted through soil by the zoospores of the chytridiomycete fungus Olpidium virulentus. During surveys carried out in two growing seasons from 2015 to 2016 in the main vegetable growing areas in Jordan (Jordan Valley, Jerash, and Madaba), 179 lettuce plants, 46 lettuce seedlings, and 56 weed plants were collected. RNA extraction was conducted from leaves of all samples. Associated viruses were identified using reverse transcription‐polymerase chain reaction (RT‐PCR) with specific primer pairs targeting part of the coat protein (CP) genes of LBVaV and MiLBVV. RT‐PCR results revealed that, out of 225 lettuce samples tested, 32 (14%) and 44 (20%) were singly infected with LBVaV and MiLBVV, respectively, and 73 (32%) samples were dually infected with LBVaV and MiLBVV. Overall, MiLBVV was detected in more lettuce samples (117) compared with LBVaV which was detected in 105 samples. Moreover, one sample of each weed species of Chenopodium murale and Sonchus oleraceus tested positive for MiLBVV and LBVaV, respectively. The partial nucleotide sequence of the CP genes of LBVaV and MiLBVV Jordanian isolates showed similarities to other isolates available in the NCBI database ranging between 93.9–100% and 88.3–100%, respectively. Nucleotide sequence comparison of the Jordanian isolates and inference from phylogenetic trees with strong support revealed well‐defined sub‐groups with little or no correlation with planting area or geographic origin. The occurrence of both viruses in lettuce plants was associated with the presence of resting spores and zoospores of Olpidium spp. in lettuce plants grown under field and growth chamber conditions. Lettuce plants inoculated with Olpidium spp. zoospores isolated from roots of symptomatic lettuce plants, developed LBVD‐symptoms. At 60 days post‐transplantation, both viruses were detected by RT‐PCR in most of the plants analysed. Biological and molecular assays together with the light microscopy observations of resting spores and zoospores of Olpidium spp., revealed the presence of LBVaV, MiLBVV, and O. virulentus in big‐vein diseased lettuce plants.